DESCRIPTION (taken from the application) The long term goals of this study are to elucidate the mechanisms of control of the hypothalamic-pituitary-thyroid (HPT) axis under basal conditions and during conditions when this regulatory system is perturbed by environmental stressors or disease states characterized by low circulating thyroid hormone and inappropriately low TSH (nonthyroidal illness syndrome). We have recently demonstrated the presence of large concentrations of type 2 iodothyronine deiodinase (D2) mRNA in a group of specialized glial cells lining the floor and infralateral wall of the third ventricle, termed D2 tanycytes. Since D2 converts thyroxine (T4) to the more biologically active thyroid hormone, triiodothyronine (T3), we propose that D2 tanycytes comprise part of a previously unrecognized regulatory component of the HPT axis. We further propose that via interactions with the cerebrospinal fluid and neurons in the adjacent hypothalamic arcuate nucleus, D2 tanycytes modulate the set point for feedback regulation of thyroid hormone on the gene expression of TRH in the hypothalamic paraventricular nucleus. The purpose of this R21 Pilot and Feasibility Proposal is to develop animal models to test the importance of the D2 tanycyte on the regulation of the HPT axis and begin to elucidate the mechanisms by which this regulation occurs. Two different approaches to developing animal models with D2 deficiency or D2 overexpression in tanycytes will be used. In the first model, adenovirus that express antisense or overexpress sense D2 mRNA will be used to infect tanycytes after stereotaxic injection of the virus into the third ventricle or median eminence. The second model will involve the development of transgenic mice, using the DARPP-32 gene as a tool to target antisense or sense D2 transcripts to the tanycytes. Efficacy of expression of the D2 antisense and sense transcripts in tanycytes will be determined by measuring D2 activity in extracts from the mediobasal hypothalamus, and the specificity for tanycytes demonstrated by in situ hybridization histochemistry. The effect of D2 depletion and overexpression in tanycytes on the HPT axis will then be studied under basal conditions or in hypothyroid and fasting animals. Due to the coexistence of DARPP-32, a phosphatase inhibitor, and D2 in tanycytes, we will also determine whether DARPP-32 exerts a regulatory function on D2 by measuring circulating thyroid hormone levels and D2 activity in tanycytes in response to hypothyroidism and fasting in the DARPP-32 knock-out mouse.